Application of Transcription Factor Decoys for Avermectin and Lycopene Overproduction in <i>Streptomyces avermitilis</i>

Here, we applied a transcription factor (TF) decoy strategy to enhance avermectin (AVE) production and activate the cryptic lycopene biosynthetic gene cluster (BGC) in industrial species <i>Streptomyces avermitilis</i>. Two decoy systems based on multicopy plasmid pKC1139 for repressors AfsR and AveI significantly increased AVE titers by 526.1% and 417.6%, respectively, compared to the wild-type (WT) strain. We further constructed a series of integrative decoy plasmids harboring varying numbers of AfsR decoys to demonstrate the dose-dependent effect, followed by a pKC1139-based AfsR decoy library to screen for optimal AVE-producing sequences. Three top-performing sequences were identified, increasing AVE titers by up to 1040.2% in the WT strain and 36.1% in an industrial strain. Finally, we successfully activated lycopene production (45.5 μg/mL) by using the <i>crtY-crtE</i> intergenic region as a decoy. These findings demonstrate that the simple, easy-to-implement TF decoy strategy can be applied to <i>Streptomyces</i> metabolic engineering for the overproduction of secondary metabolites.